Hi everyone, I was trying to check the RT-PCR product which is less than 100bp on the agarose gel. I used 3% agarose and ran 80V for 1.5 hr, then I put my gel in the 1XTAE buffer with GelRED to stain about 1hr. I could see my low molecular ladder, however, it is so hard to see the band of PCR products. Should I change to another agarose? Does anyone have a suggestion for that? Thank you so much!
It is a very small product so probably separates better on PAGE but if you are using agarose then you should run the gel as cool as possible at a low voltage and for a short time. The low voltage means less heating so less diffusion of the band and the short running time means the band is at its most concentrated so it should show up better. The percentage is fine at 3% but try running for 30 minutes and loading twice as much pcr product. You do not need all of the larger ladder bands to show just the bottom 2 will do
Paul Rutland Thank you so much for your suggestion, Paul! It makes sense. I will try to run my gel faster using low voltage.
Chen Zhang do not worry about the run being too short. You can always put the gel back in the gel tank to run it further if you think it is needed
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